1. Field of the Invention
The present invention relates to analytical chemistry, and, more particularly, to devices and methods for determining the concentration of analytes in liquid samples.
2. State of the Art
In the field of analytical chemistry, there is a continuing interest in developing new, simpler and more reliable techniques to detect and measure the presence of analyte(s) in samples. In many instances, both speed and accuracy are important for the measurement, particularly with certain physiologically active compounds. In other situations, convenience can be made a major consideration.
Piezoelectric sensors have been used as microgravimetric immunoassay devices (see, for example, Joy E. Roederer and Glenn J. Bastiaans, "Microgravimetric Immunoassay with Piezoelectric Crystals", Anal. Chem. 1983, 2333-2336). These sensors operate on the principle that changes in the amount of mass attached to their surface cause shifts in the resonant frequency.
For ascertaining the concentration of a compound in a liquid, the piezoelectric sensor is preferably a surface transverse wave device or a Love wave device (collectively referred to as "piezoelectric surface wave devices"). Such devices are known in the art and are disclosed, for example, in U.S. Pat. No. 5,130,257 and U.S. patent application Ser. Nos. 07/792,975 and 07/404,721, the disclosures of which are incorporated herein by reference in their entirety.
Piezoelectric surface wave devices and other surface acoustic wave devices have come into general use, primarily for filtering radio frequency signals. A typical device is constructed on a piezoelectric substrate and has interdigital input and output transducers defined by precise electrode fingers.
In the case of piezoelectric surface wave devices, selective mass detection is achieved by coating the surface of the device with a chemically reactive layer (e.g., a receptor layer) that preferentially reacts with the substance to be detected such that the mass of the receptor layer changes in the presence of this substance. These devices function as chemical sensors that can measure the concentration of the selected class of compounds in a solution into which the sensor or device is immersed. For example, to measure the concentration of a specific antibody in a solution, a piezoelectric surface wave device is utilized in which the receptor layer contains the antigen corresponding to this antibody. The concentration of this antibody in a liquid can be measured by immersing the device in the liquid. Antibody present in the liquid will bind to the surface of the device, thereby increasing the mass loading of the top surface. Radio frequency energy coupled into the device through the input transducer is converted to a surface acoustic wave confined to within a few wavelengths of the surface. The velocity of the surface acoustic wave will vary according to the mass loading of the top surface of the device. The surface acoustic wave propagates along the surface of the device until it encounters the output transducer, which converts the surface acoustic wave back into RF energy. The change in velocity of the surface acoustic wave with the mass loading of the surface of the device translates into variation of the phase of the radio frequency signal output by the output transducer which can then be correlated to the mass of the antibody bound to the surface.
The mass sensitivity (i.e., the fractional frequency change divided by the mass change of material deposited on the surface of the device) increases as the mass of the piezoelectric surface wave device is decreased or, correspondingly, as the device thickness is decreased. A practical lower limit of about 100 microns, corresponding to a resonance frequency of about 20 MHz, is imposed on device thickness by manufacturing difficulties. Consequently, the sensitivity of piezoelectric surface wave devices is limited.
As an additional complication, while prior art devices attribute the shift in resonant frequency in piezoelectric surface wave devices used in liquid sample (e.g., aqueous solutions) to changes in the mass attached to the surface, not all of this shift is due solely to increases in the mass. In particular, physical and chemical factors such as temperature, pressure, non-specific binding, ionic strength, conductivity, mass density, viscosity, etc., are all interrelated to the shift in resonant frequency and interfere with any direct correlation between the shift in resonant frequency and the changes in mass attached to the surface of the device. While this interference can be minimized to some extent, a true correlation between frequency shifts and mass is not possible.
In the prior art, multiple measurements of multiple analytes have required a large volume of analyte to be divided into separate samples, one for each measurement. In certain medical testing applications, only a limited quantity of analyte may be available. Without the ability to perform multiple measurements of multiple analytes from a single sample, the number of measurements that may be performed is therefore limited.